INTERNALIZATION OF HYALURONAN BY CHONDROCYTES OCCURS VIA RECEPTOR-MEDIATED ENDOCYTOSIS

Several studies have suggested that chondrocytes must have the capacit y to internalize and degrade extracellular hyaluronan. In the present study we show direct evidence that hyaluronan is, in fact, endocytosed by chondrocytes and that the endocytosis is mediated via cell surface CD44/hyaluronan receptors. Cultures of bovine articular chondrocytes as well as rat chondrosarcoma chondrocytes were incubated with either fluorescein- or 3H-labeled hyaluronan. Intense binding and accumulatio n of labeled hyaluronan was visualized by fluorescence microscopy or b right-field/dark-field microscopy following autoradiography. Cell surf ace hyaluronan was removed with either trypsin or Streptomyces hyaluro nidase in order to distinguish and quantify intracellular endocytosed hyaluronan. Labeled hyaluronan was visualized within small discrete in tracellular vesicles distributed throughout the cytoplasm. Binding and endocytosis of fluorescein- or H-3-labeled hyaluronan was totally blo cked by the addition of excess unlabeled hyaluronan or hyaluronan hexa saccharides, competitive inhibitors of hyaluronan/hyaluronan receptor interactions. Binding and endocytosis was also blocked by the addition of anti-CD44 monoclonal antibodies. Characterization of endocytosed H -3-labeled hyaluronan demonstrated that a significant portion of the h yaluronan was degraded by both the bovine articular and rat chondrosar coma chondrocytes. Interestingly, a higher proportion of bound hyaluro nan was internalized by the bovine chondrocytes. Therefore, hyaluronan receptor-mediated endocytosis and degradation of hyaluronan may provi de a critical link to the maintenance and homeostasis of cartilage tis sue.